Study On The Regulation Mechanism Of 4-Hydroxybenzoic Acid On The Secretory Properties Of Grapevine Roots

Autotoxicity is the key reason of the occurence of grape replant disease.The previous research is found that 4-hydroxybenzoic acid（4-HBA）as an autotoxic substance has a certain regulatory effect on the secretory properties of grape roots,but the specific signal transduction pathway involved in regulation is not yet known.Therefore,in this study,we used grape rootstock’Beta’as the test material,4-HBA as the toxic substances,and we set different concentration levels and sampling time.We determined the best concentration and time of 4-HBA stress treatment by measuring the root defense enzyme SOD activity,phenylalanine ammonia-lyase（PAL）activity,malondialdehyde（MDA）and root phenolic acid content changes in Beta grape roots.Then,different signal transduction inhibitors are co-treated with4-HBA to clarify the signal transduction pathways involved in the process of 4-HBA acid-induced changes in the secretory properties of Beta grape roots.1.Exogenous 4-HBA treatment affected the growth,SOD activity,PAL activity,and MDA content of Beta grape seedlings.250μmol·L^-1,500μmol·L^-1,1000μmol·L^-11 4-HBA can inhibit the growth of Beta seedlings,and the inhibitory effect is more pronounced with the increase of 4-HBA concentration,the effect in fresh weight of shoot and root is stronger than plant height.The activities of SOD enzyme and PAL enzyme showed a trend of increasing first and then decreasing with the prolonging of treatment time,and all peaked at 48h,and the MDA content continued to accumulate.At the early stage of stress,there was no significant difference in the activity of SOD enzyme under the treatment of exogenous 4-HBA at various concentrations.In the late period of stress,low concentration of phenolic acid promoted the activity of SOD enzyme and inhibited its activity at high concentration.PAL enzyme activity increased first and then decreased with increasing treatment concentration.2.Exogenous 4-HBA treatment affected the phenolic acid content in root exudates and root tissue.Exogenous 4-HBA treatment resulted in a decrease in the content of 4-HBA excreted in roots,salicylic acid was significantly increased,and the total phenolic acid content decreased at high concentrations at high concentrations,which was significantly different from the control.Treatment with exogenous 4-HBA increased the content of 4-HBA and salicylic acid in the root tissue,which was significantly different from the control,and increased with increasing treatment concentration.3.Signal transduction inhibitor treatment affects the growth of tissue culture seedlings of Beta grape.Both 25μmol·L^-1 and 150μmol·L^-11 VP could relieve the stress of 4-HBA on the growth of tissue culture seedlings,and the effect of high concentration stress relief was more significant.1μmol·L^-1 DPI could relieve the stress of 4-HBA on the growth of tissue culture seedlings,while high concentration of DPI had no significant effect.DETC of 1μmol·L^-1 and10μmol·L^-1 could alleviate the stress of 4-HBA on the growth of tissue culture seedlings,and the 10μmol·L^-1 DETC treatment had a more significant effect.4.The treatment of exogenous phenolic acid at the root tip of Bettegrain resulted in a significant change in intracellular Ca²⁺distribution,which gradually spread from the cell wall to the cytoplasm.Treatment with Ca²⁺channel inhibitors VP and LaCl₃ reduced the concentration of Ca²⁺in the cytoplasm.The effect of LaCl₃ was even more pronounced at the same concentration of 150μmol·L^-1.5.Calcium channel inhibitors,NADPH oxidase inhibitors,and SOD specific inhibitors can inhibit the induction of phenolic acids from roots by 4-HBA.Treatments of Vp,LaCl₃ and4-HBA together resulted in a significant decrease in PAL activity and 4-HBA and salicylic acid content in root exudates.It is suggested that the extracellular calcium ion induced by 4-HBA is involved in the signal transduction process of phenolic acid biosynthesis in grape roots.The combination of DETC and 4-HBA treatment resulted in decreased PAL activity in grape roots and decreased 4-HBA content in root exudates.After treatment with DPI and 4-HBA,there was no significant change in PAL activity.